Protein secretion is a fundamentally conserved process that provides a mechanism for cells to create stable intracellular compartments such as the ER, Golgi, lysosome, and plasma membrane, and to communicate with external cells and tissues. A genetic, molecular cloning, and biochemical analysis of this process in Saccharomyces cerevisiae has uncovered genes and mechanisms that are shared among all eukaryotes. This proposal develops a new line of investigation concerning the mechanism of transport vesicle formation from the trans Golgi cisterna and of the pathway of vesicle traffic to the cytokinesis furrow which forms early in the yeast cell cycle. Evidence has emerged to suggest multiple paths of transport vesicle formation from the trans Golgi, at least one of which involves vesicular traffic through the endosome en route to the cell surface. Genetic and cell fractionation analysis is proposed to relate secretion to the known pathway of sorting to the endosome, to discover new genes involved in budding from the Golgi, and to use biochemical analysis to clarify the role of clathrin in one of the budding pathways. Certain membrane enzymes are transported through the endosome to the nascent division septum where they participate in forming a cell wall structure that marks the site of yeast bud emergence (the chitin ring) early in the cell cycle. The pathway and regulation of transport of these enzymes from the endosome to the nascent septeum will be examined to understand the mechanism and cell cycle control of this essential process. The study of these processes in yeast cells has illuminated fundamental aspects of cell and organelle growth that are shared by human cells. Furthermore, the application of this knowledge has allowed biotechnology and pharmaceutical companies to harness yeast cells to express and secrete commercial quantities of important hormones, blood proteins, growth factors, and viral antigens.